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bioRxiv · 10.1101/2023.06.21.545902

Kinetic and functional analysis of abundant microRNAs in extracellular vesicles from normal and stressed cultures of Chinese Hamster Ovary (CHO) cells

Belliveau, J.·Thompson, W.·Papoutsakis, E. T.
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Abstract

Chinese hamster ovary (CHO) cells release and exchange large quantities of extracellular vesicles (EVs). EVs are highly enriched in microRNAs (miRs, or miRNAs), which are responsible for most of their biological effects. We have recently shown that the miR content of CHO EVs varies significantly under culture stress conditions. Here, we provide a novel stoichiometric ("per-EV") quantification of miR and protein levels in large CHO EVs produced under ammonia, lactate, osmotic, and age-related stress. Each stress resulted in distinct EV miR levels, with selective miR loading by parent cells. Our data provide a proof of concept for the use of CHO EV cargo as a diagnostic tool for identifying culture stress. We also tested the impact of three select miRs (let-7a, miR-21, and miR-92a) on CHO cell growth and viability. Let-7a--abundant in CHO EVs from stressed cultures--reduced CHO cell viability, while miR-92a--abundant in CHO EVs from unstressed cultures--promoted cell survival. Overexpression of miR-21 had a slight detrimental impact on CHO cell growth and viability during late exponential-phase culture, an unexpected result based on the reported anti-apoptotic role of miR-21 in other mammalian cell lines. These findings provide novel relationships between CHO EV cargo and cell phenotype, suggesting that CHO EVs may exert both pro- and anti-apoptotic effects on target cells, depending on the conditions under which they were produced. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=91 SRC="FIGDIR/small/545902v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1f85cd7org.highwire.dtl.DTLVardef@973599org.highwire.dtl.DTLVardef@13ff81eorg.highwire.dtl.DTLVardef@ba8eba_HPS_FORMAT_FIGEXP M_FIG C_FIG

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Belliveau, J., Thompson, W., Papoutsakis, E. T.. 2023-06-24. Kinetic and functional analysis of abundant microRNAs in extracellular vesicles from normal and stressed cultures of Chinese Hamster Ovary (CHO) cells. https://doi.org/10.1101/2023.06.21.545902

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